bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2021‒08‒15
eighteen papers selected by
Ralitsa Radostinova Madsen
University College London Cancer Institute

  1. Proc Natl Acad Sci U S A. 2021 Aug 17. pii: e2101496118. [Epub ahead of print]118(33):
      The protein kinase Akt is one of the primary effectors of growth factor signaling in the cell. Akt responds specifically to the lipid second messengers phosphatidylinositol-3,4,5-trisphosphate [PI(3,4,5)P3] and phosphatidylinositol-3,4-bisphosphate [PI(3,4)P2] via its PH domain, leading to phosphorylation of its activation loop and the hydrophobic motif of its kinase domain, which are critical for activity. We have now determined the crystal structure of Akt1, revealing an autoinhibitory interface between the PH and kinase domains that is often mutated in cancer and overgrowth disorders. This interface persists even after stoichiometric phosphorylation, thereby restricting maximum Akt activity to PI(3,4,5)P3- or PI(3,4)P2-containing membranes. Our work helps to resolve the roles of lipids and phosphorylation in the activation of Akt and has wide implications for the spatiotemporal control of Akt and potentially lipid-activated kinase signaling in general.
    Keywords:  Akt; PIP3; kinase; lipid; signaling
  2. Genet Med. 2021 Aug 12.
      PURPOSE: PIK3CA pathogenic variants in the PIK3CA-related overgrowth spectrum (PROS) activate phosphoinositide 3-kinase signaling, providing a rationale for targeted therapy, but no drug has proven efficacy and safety in this population. Our aim was to establish the six-month tolerability and efficacy of low-dose taselisib, a selective class I PI3K inhibitor, in PROS patients.METHODS: Patients over 16 years with PROS and PIK3CA pathogenic variants were included in a phase IB/IIA multicenter, open-label single-arm trial (six patients at 1 mg/day of taselisib, then 24 at 2 mg/day). The primary outcome was the occurrence of dose limiting toxicity (DLT). Efficacy outcomes were the relative changes after treatment of (1) tissue volume at affected and unaffected sites, both clinically and on imaging; (2) cutaneous vascular outcomes when relevant; (3) biologic parameters; (4) quality of life; and (5) patient-reported outcomes.
    RESULTS: Among 19 enrolled patients, 2 experienced a DLT (enteritis and pachymeningitis) leading to early trial termination (17 treated, 10 completed the study). No serious adverse reaction occurred in the 1 mg cohort (n = 6). No significant reduction in affected tissue volume was observed (mean -4.2%; p = 0.81; SD 14.01). Thirteen (76.4%) participants reported clinical improvement (pain reduction, chronic bleeding resolution, functional improvement).
    CONCLUSION: Despite functional improvement, the safety profile of low-dose taselisib precludes its long-term use.
  3. Nat Commun. 2021 08 10. 12(1): 4814
      Glutamoptosis is the induction of apoptotic cell death as a consequence of the aberrant activation of glutaminolysis and mTORC1 signaling during nutritional imbalance in proliferating cells. The role of the bioenergetic sensor AMPK during glutamoptosis is not defined yet. Here, we show that AMPK reactivation blocks both the glutamine-dependent activation of mTORC1 and glutamoptosis in vitro and in vivo. We also show that glutamine is used for asparagine synthesis and the GABA shunt to produce ATP and to inhibit AMPK, independently of glutaminolysis. Overall, our results indicate that glutamine metabolism is connected with mTORC1 activation through two parallel pathways: an acute alpha-ketoglutarate-dependent pathway; and a secondary ATP/AMPK-dependent pathway. This dual metabolic connection between glutamine and mTORC1 must be considered for the future design of therapeutic strategies to prevent cell growth in diseases such as cancer.
  4. Sci Rep. 2021 Aug 11. 11(1): 16344
      Placenta growth factor (PlGF) is a pro-inflammatory angiogenic mediator that promotes many pathologies including diabetic complications and atherosclerosis. Widespread endothelial dysfunction precedes the onset of these conditions. As very little is known of the mechanism(s) controlling PlGF expression in pathology we investigated the role of hyperglycaemia in the regulation of PlGF production in endothelial cells. Hyperglycaemia stimulated PlGF secretion in cultured primary endothelial cells, which was suppressed by IGF-1-mediated PI3K/Akt activation. Inhibition of PI3K activity resulted in significant PlGF mRNA up-regulation and protein secretion. Similarly, loss or inhibition of Akt activity significantly increased basal PlGF expression and prevented any further PlGF secretion in hyperglycaemia. Conversely, constitutive Akt activation blocked PlGF secretion irrespective of upstream PI3K activity demonstrating that Akt is a central regulator of PlGF expression. Knock-down of the Forkhead box O-1 (FOXO1) transcription factor, which is negatively regulated by Akt, suppressed both basal and hyperglycaemia-induced PlGF secretion, whilst FOXO1 gain-of-function up-regulated PlGF in vitro and in vivo. FOXO1 association to a FOXO binding sequence identified in the PlGF promoter also increased in hyperglycaemia. This study identifies the PI3K/Akt/FOXO1 signalling axis as a key regulator of PlGF expression and unifying pathway by which PlGF may contribute to common disorders characterised by endothelial dysfunction, providing a target for therapy.
  5. Nat Commun. 2021 08 10. 12(1): 4838
      Macropinosomes are formed by shaping actin-rich plasma membrane ruffles into large intracellular organelles in a phosphatidylinositol 3-kinase (PI3K)-coordinated manner. Here, we utilize lattice lightsheet microscopy and image visualization methods to map the three-dimensional structure and dynamics of macropinosome formation relative to PI3K activity. We show that multiple ruffling morphologies produce macropinosomes and that the majority form through collisions of adjacent PI3K-rich ruffles. By combining multiple volumetric representations of the plasma membrane structure and PI3K products, we show that PI3K activity begins early throughout the entire ruffle volume and continues to increase until peak activity concentrates at the base of the ruffle after the macropinosome closes. Additionally, areas of the plasma membrane rich in ruffling had increased PI3K activity and produced many macropinosomes of various sizes. Pharmacologic inhibition of PI3K activity had little effect on the rate and morphology of membrane ruffling, demonstrating that early production of 3'-phosphoinositides within ruffles plays a minor role in regulating their morphology. However, 3'-phosphoinositides are critical for the fusogenic activity that seals ruffles into macropinosomes. Taken together, these data indicate that local PI3K activity is amplified in ruffles and serves as a priming mechanism for closure and sealing of ruffles into macropinosomes.
  6. Nature. 2021 Aug 11.
      Non-genetic mechanisms have recently emerged as important drivers of cancer therapy failure1, where some cancer cells can enter a reversible drug-tolerant persister state in response to treatment2. Although most cancer persisters remain arrested in the presence of the drug, a rare subset can re-enter the cell cycle under constitutive drug treatment. Little is known about the non-genetic mechanisms that enable cancer persisters to maintain proliferative capacity in the presence of drugs. To study this rare, transiently resistant, proliferative persister population, we developed Watermelon, a high-complexity expressed barcode lentiviral library for simultaneous tracing of each cell's clonal origin and proliferative and transcriptional states. Here we show that cycling and non-cycling persisters arise from different cell lineages with distinct transcriptional and metabolic programs. Upregulation of antioxidant gene programs and a metabolic shift to fatty acid oxidation are associated with persister proliferative capacity across multiple cancer types. Impeding oxidative stress or metabolic reprogramming alters the fraction of cycling persisters. In human tumours, programs associated with cycling persisters are induced in minimal residual disease in response to multiple targeted therapies. The Watermelon system enabled the identification of rare persister lineages that are preferentially poised to proliferate under drug pressure, thus exposing new vulnerabilities that can be targeted to delay or even prevent disease recurrence.
  7. J Cell Sci. 2022 Mar 01. pii: jcs258364. [Epub ahead of print]135(5):
      The lipid composition of the primary cilia membrane is emerging as a critical regulator of cilia formation, maintenance and function. Here, we show that conditional deletion of the phosphoinositide 5'-phosphatase gene Inpp5e, mutation of which is causative of Joubert syndrome, in terminally developed mouse olfactory sensory neurons (OSNs), leads to a dramatic remodeling of ciliary phospholipids that is accompanied by marked elongation of cilia. Phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2], which is normally restricted to the proximal segment redistributed to the entire length of cilia in Inpp5e knockout mice with a reduction in phosphatidylinositol (3,4)-bisphosphate [PI(3,4)P2] and elevation of phosphatidylinositol (3,4,5)-trisphosphate [PI(3,4,5)P3] in the dendritic knob. The redistribution of phosphoinositides impaired odor adaptation, resulting in less efficient recovery and altered inactivation kinetics of the odor-evoked electrical response and the odor-induced elevation of cytoplasmic Ca2+. Gene replacement of Inpp5e through adenoviral expression restored the ciliary localization of PI(4,5)P2 and odor response kinetics in OSNs. Our findings support the role of phosphoinositides as a modulator of the odor response and in ciliary biology of native multi-ciliated OSNs.
    Keywords:  INPP5E; Mouse; Odor response; Olfactory cilia; Phospholipids
  8. Proc Natl Acad Sci U S A. 2021 Aug 17. pii: e2111313118. [Epub ahead of print]118(33):
  9. Proc Natl Acad Sci U S A. 2021 Aug 17. pii: e2103261118. [Epub ahead of print]118(33):
      RAS proteins are molecular switches that interact with effector proteins when bound to guanosine triphosphate, stimulating downstream signaling in response to multiple stimuli. Although several canonical downstream effectors have been extensively studied and tested as potential targets for RAS-driven cancers, many of these remain poorly characterized. In this study, we undertook a biochemical and structural approach to further study the role of Sin1 as a RAS effector. Sin1 interacted predominantly with KRAS isoform 4A in cells through an atypical RAS-binding domain that we have characterized by X-ray crystallography. Despite the essential role of Sin1 in the assembly and activity of mTORC2, we find that the interaction with RAS is not required for these functions. Cells and mice expressing a mutant of Sin1 that is unable to bind RAS are proficient for activation and assembly of mTORC2. Our results suggest that Sin1 is a bona fide RAS effector that regulates downstream signaling in an mTORC2-independent manner.
    Keywords:  KRAS; RAS; RBD; Sin1; mTORC2
  10. Stem Cell Reports. 2021 Jul 29. pii: S2213-6711(21)00380-5. [Epub ahead of print]
      The health and homeostasis of skeletal muscle are preserved by a population of tissue-resident muscle stem cells (MuSCs) that maintain a state of mitotic and metabolic quiescence in adult tissues. The capacity of MuSCs to preserve the quiescent state declines with aging and metabolic insults, promoting premature activation and stem cell exhaustion. Sestrins are a class of stress-inducible proteins that act as antioxidants and inhibit the activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling complex. Despite these pivotal roles, the role of Sestrins has not been explored in adult stem cells. We show that SESTRIN1,2 loss results in hyperactivation of the mTORC1 complex, increased propensity to enter the cell cycle, and shifts in metabolic flux. Aged SESTRIN1,2 knockout mice exhibited loss of MuSCs and a reduced ability to regenerate injured muscle. These findings demonstrate that Sestrins help maintain metabolic pathways in MuSCs that protect quiescence against aging.
    Keywords:  RNA sequencing; aging; mTORC1; metabolism; oxidative stress; reactive oxygen species; regeneration; satellite cells
  11. Sci Signal. 2021 Aug 10. pii: eabc0836. [Epub ahead of print]14(695):
      Exogenous administration of lymphangiogenic growth factors is widely used to study changes in lymphatic function in pathophysiology. However, this approach can result in off-target effects, thereby generating conflicting data. To circumvent this issue, we modulated intracellular VEGF-C signaling by conditionally knocking out the lipid phosphatase PTEN using the Vegfr3 promoter to drive the expression of Cre-lox in lymphatic endothelial cells (LECs). PTEN is an intracellular brake that inhibits the downstream effects of the activation of VEGFR3 by VEGF-C. Activation of Cre-lox recombination in adult mice resulted in an expanded functional lymphatic network due to LEC proliferation that was independent of lymphangiogenic growth factor production. Furthermore, compared with lymphangiogenesis induced by VEGF-C injection, LECPTEN animals had mature, nonleaky lymphatics with intact cell-cell junctions and reduced local tissue inflammation. Last, compared with wild-type or VEGF-C-injected mice, LECPTEN animals had an improved capacity to resolve inflammatory responses. Our findings indicate that intracellular modulation of lymphangiogenesis is effective in inducing functional lymphatic networks and has no off-target inflammatory effects.
  12. Immunity. 2021 Aug 10. pii: S1074-7613(21)00295-8. [Epub ahead of print]54(8): 1807-1824.e14
      The transcription factor forkhead box O1 (FOXO1), which instructs the dark zone program to direct germinal center (GC) polarity, is typically inactivated by phosphatidylinositol 3-kinase (PI3K) signals. Here, we investigated how FOXO1 mutations targeting this regulatory axis in GC-derived B cell non-Hodgkin lymphomas (B-NHLs) contribute to lymphomagenesis. Examination of primary B-NHL tissues revealed that FOXO1 mutations and PI3K pathway activity were not directly correlated. Human B cell lines bearing FOXO1 mutations exhibited hyperactivation of PI3K and Stress-activated protein kinase (SAPK)/Jun amino-terminal kinase (JNK) signaling, and increased cell survival under stress conditions as a result of alterations in FOXO1 transcriptional affinities and activation of transcriptional programs characteristic of GC-positive selection. When modeled in mice, FOXO1 mutations conferred competitive advantage to B cells in response to key T-dependent immune signals, disrupting GC homeostasis. FOXO1 mutant transcriptional signatures were prevalent in human B-NHL and predicted poor clinical outcomes. Thus, rather than enforcing FOXO1 constitutive activity, FOXO1 mutations enable co-option of GC-positive selection programs during the pathogenesis of GC-derived lymphomas.
    Keywords:  B cell; CD40; FOXO1; JNK; PI3K; germinal center; mouse model; mutation; non-Hodgkin lymphoma; positive selection
  13. Cell Metab. 2021 Aug 03. pii: S1550-4131(21)00332-6. [Epub ahead of print]
      Cancer cells are metabolically similar to their corresponding normal tissues. Differences between cancers and normal tissues may reflect reprogramming during transformation or maintenance of the metabolism of the specific normal cell type that originated the cancer. Here, we compare glucose metabolism in hematopoiesis and leukemia. Thymus T cell progenitors were glucose avid and oxidized more glucose in the tricarboxylic acid cycle through pyruvate dehydrogenase (PDH) as compared with other hematopoietic cells. PDH deletion decreased double-positive T cell progenitor cells but had no effect on hematopoietic stem cells, myeloid progenitors, or other hematopoietic cells. PDH deletion blocked the development of Pten-deficient T cell leukemia, but not the development of a Pten-deficient myeloid neoplasm. Therefore, the requirement for PDH in leukemia reflected the metabolism of the normal cell of origin independently of the driver genetic lesion. PDH was required to prevent pyruvate accumulation and maintain glutathione levels and redox homeostasis.
    Keywords:  T cell leukemia; double-positive thymocytes; glycolysis; hematopoietic stem cells; metabolism; pyruvate dehydrogenase; thymus
  14. Cancer Cell. 2021 Aug 09. pii: S1535-6108(21)00390-1. [Epub ahead of print]39(8): 1047-1049
      Resistance to targeted therapies is a major challenge in cancer care and occurs via genetic and non-genetic mechanisms. In this issue of Cancer Cell, Marin-Bejar et al. demonstrate that melanomas recurrently select genetic or non-genetic resistance trajectories and that targeting neural crest stem cell-like cells prevents non-genetic, but not genetic, resistance.
  15. Mol Cancer Res. 2021 Aug 13. pii: molcanres.MCR-21-0060-A.2021. [Epub ahead of print]
      Previous studies have reported dysregulated cytoplasmic and nuclear expression of the β-catenin protein in triple-negative breast cancer (TNBC) in the absence of Wnt signaling pathway dysregulation. However, the mechanism that sustains β-catenin protein dysregulation independent of Wnt signaling is not understood. In this study, we show that SHP2 is essential for β-catenin protein stability and for sustaining the cytoplasmic and nuclear pools in TNBC cells. The first evidence for this possibility came from immunofluorescence (IF) and immunoblotting (IB) studies that showed that inhibition of SHP2 induces E-cadherin expression and depletion of cytoplasmic and nuclear β-catenin, which in turn confers adherence junction mediated cell-cell adhesion. We further show that SHP2 promotes β-catenin protein stability by mediating the inactivation of GSK3β through its positive effect on Akt and ERK1/2 activation, which was confirmed by direct pharmacological inhibition of the PI3K-Akt and the MEK-ERK signaling pathway. Finally, we show that SHP2-stabilized β-catenin contributes to TNBC cell growth, transformation, CSC properties, and tumorigenesis and metastasis. Overall, the findings in this report show that SHP2 mediates β-catenin protein stability to promote TNBC. Implications: Data presented in this article demonstrates that SHP2 positively regulates β-catenin protein stability, which in turn promotes triple-negative breast cancer cell transformation, tumorigenesis, and metastasis.
  16. Nucleic Acids Res. 2021 Aug 07. pii: gkab686. [Epub ahead of print]
      CRISPR-Cas9 generates double-stranded DNA breaks (DSBs) to activate cellular DNA repair pathways for genome editing. The repair of DSBs leads to small insertions or deletions (indels) and other complex byproducts, including large deletions and chromosomal translocations. Indels are well understood to disrupt target genes, while the other deleterious byproducts remain elusive. We developed a new in silico analysis pipeline for the previously described primer-extension-mediated sequencing assay to comprehensively characterize CRISPR-Cas9-induced DSB repair outcomes in human or mouse cells. We identified tremendous deleterious DSB repair byproducts of CRISPR-Cas9 editing, including large deletions, vector integrations, and chromosomal translocations. We further elucidated the important roles of microhomology, chromosomal interaction, recurrent DSBs, and DSB repair pathways in the generation of these byproducts. Our findings provide an extra dimension for genome editing safety besides off-targets. And caution should be exercised to avoid not only off-target damages but also deleterious DSB repair byproducts during genome editing.
  17. Nat Commun. 2021 Aug 13. 12(1): 4920
      Malignant mesothelioma (MpM) is an aggressive, invariably fatal tumour that is causally linked with asbestos exposure. The disease primarily results from loss of tumour suppressor gene function and there are no 'druggable' driver oncogenes associated with MpM. To identify opportunities for management of this disease we have carried out polysome profiling to define the MpM translatome. We show that in MpM there is a selective increase in the translation of mRNAs encoding proteins required for ribosome assembly and mitochondrial biogenesis. This results in an enhanced rate of mRNA translation, abnormal mitochondrial morphology and oxygen consumption, and a reprogramming of metabolic outputs. These alterations delimit the cellular capacity for protein biosynthesis, accelerate growth and drive disease progression. Importantly, we show that inhibition of mRNA translation, particularly through combined pharmacological targeting of mTORC1 and 2, reverses these changes and inhibits malignant cell growth in vitro and in ex-vivo tumour tissue from patients with end-stage disease. Critically, we show that these pharmacological interventions prolong survival in animal models of asbestos-induced mesothelioma, providing the basis for a targeted, viable therapeutic option for patients with this incurable disease.
  18. J Cell Sci. 2021 Aug 12. pii: jcs.258428. [Epub ahead of print]
      Animals subjected to dietary restriction (DR) have reduced body size, low fecundity, slower development, lower fat content and longer life span. We identified lamin as a regulator of multiple dietary restriction phenotypes. Downregulation of lmn-1, the single Caenorhabditis elegans lamin gene, increased animal size and fat content, specifically in DR animals. The LMN-1 protein acts in the mTOR pathway, upstream to RAPTOR and S6K, key component and target of mTOR complex 1 (mTORC1), respectively. DR excludes the mTORC1 activator RAGC-1 from the nucleus. Downregulation of lmn-1 restores RAGC-1 to the nucleus, a necessary step for the activation of the mTOR pathway. These findings further link lamin to metabolic regulation.
    Keywords:  Caenorhabditis elegans; Dietary restriction; Lamin; mTOR